The main objective of this proposal is to define the clinical relevance of detecting circulating tumor cells (CTC) in bladder cancer patients. The specific aims are: 1) To compare the detection rates of CTC for uroplakins (I-Ill), keratins (k19 and k20), and Epidermal Growth Factor Receptor (EGFR) in patients with superficial bladder cancer versus those with metastatic bladder cancer, and 2) To examine the relation between the expression of these molecular markers in primary and metastatic tissue deposits and their CTC detection rate. We propose to test blood samples from 50 bladder cancer patients with superficial tumors (expected to have low likelihood of micrometastases) and 50 with grossly metastatic disease (expected to have a high likelihood of micrometastases) using Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR). All patients will be sampled three times over a four week period to increase the positive CTC detection rate. The CTC RT-PCR will be then correlated with presence of disease at the time of the assay to define the most clinically informative marker(s) that provide the best sensitivity, specificity and overall predictive value for the presence of micrometastases. The tissue expression of the molecular markers will be assessed using immunohistochemistry as well as RT-PCR in specimens obtained from patients outlined in aim 1, and the results will then be correlated with the RT-PCR results for CTC. This analysis will help us to understand changes in the tissue expression of these markers during the progression of bladder cancer and how these changes are related to seeding of CTC in the blood. Our preliminary data clearly demonstrate the feasibility of conducting the proposed work as a collaborative effort between New York University School of Medicine and Memorial Sloan-Kettering Cancer Center. This cooperative work endows the study with the strong laboratory and clinical facilities that are necessary for the successful conduct of this type of translation research. We plan this project as a phase I pilot study, in response to the "Pilot and Feasibility Program in Urology" Program Announcement #02-013; we intend to generate sufficient data in order to pursue the development of CTC in a phase II prospective study that will examine the correlation between CTC detection and recurrence in patients with muscle invasive disease following surgical resection of the tumor. We foresee phase III marker development as a multi-institutional study with the goals of independently verifying and validating this correlation prior to considering an application to the FDA for integrating CTC detection as part of the standard of care management for bladder cancer patients.